血管内皮细胞生长因子体外转染血管内皮祖细胞的研究

Study of VEGF gene transfection on endothelial progenitor cell in vitro

目的 探讨脂质体介导血管内皮细胞生长因子(VEGF)基因转染血管内皮祖细胞(EPC)应用于基因治疗的可行性、安全性。方法 体外分离、培养、免疫组织化学及流式细胞仪鉴定EPC,转染VEGF后用免疫组织化学和ELISA检测EPC表达VEGF蛋白,噻唑蓝(MTT)检测EPC对VEGF质粒转染的敏感性。结果 EPC表达CD_(34)、CD_(31)、KDR及CD_(133)细胞表面标志,转染后EPC胞内表达VEGF。脂质体介导PcDNA3.1(-)/VEGF_(165)质粒转染、PcDNA3.1(-)/VEGF_(165)空质粒转染、不转染任何质粒的3组EPC培养基上清中表达的VEGF分别为(352±35)ng/L、(45±5)ng/L、0 ng/L(P<0.05),VEGF质粒转染对EPC增殖无影响。结论 EPC可作为VEGF转染的靶细胞,用于基因治疗。

Objective To investigate the feasibility andsecurity of VEGF transfection on endothelial progenitor cells (EPCs) mediated by liposome. Methods EPCs were isolated from human cord blood, cultured in vitro, identified by immunocytochemistry and fluorescence-activated cell sorting. After transfection, the expression of VEGF was detected by using immunocytochemistry and ELISA. MTT method was used to study the effect of transfection by VEGF on EPCs. Results EPCs expressed cell markers CD_(34), CD_(31), x 10~3R and CD_(133). After transfection the expression of VEGF was positive. The expression of VEGF in EPC's medium of three groupswhich were EPCs transfected with PcDNA3.1(-)/VEGF_(165) by liposome, EPCs transfected with nake PcDNA3.1 (-)/VEGF_(165) and EPCs without transfection was (352±35), (45±5) and 0 ng/L ( P<0.05 ). VEGF transfection had no significantly inhibitory effects on EPCs. Conclusion EPC can he used as the target cell for VEGF transfection.